The proposed studies comprise an investigation of the ability of Gram-negative bacteria to defend themselves against the spread of ssRNA bacteriophage infection. In populations of animal cells, virus-infected cells secrete interferons, polypeptides that stimulate nearby uninfected cells to activate and, in some instances, to secrete ribonucleases that confer on those cells a temporary anti-viral state. A similar phenomenon has been observed in populations of Caulobacter crescentus infected with the ssRNA phage OCb5. The first purpose of the proposed experiments is to characterize the secreted anti-viral ribonuclease by concentrating and purifying the enzyme from supernatants of infected culture and determining its substrate specificity, products, and molecular size. On the initial assumption that the signal substance that diffuses from infected cells (tentatively, "bacterial interferon, bIFN") is a polypeptide, the protein purification methods used to prepare the ribonuclease will also be used to prepare the bIFN; the assays for bIFN will test for stimulation of phage inactivation and RNase secretion by phage-resistant mutants. In addition to the protective ribonuclease that restricts the spread of infection through the population, the mildness of OCb5 infection is seen in the fate of infected cells. As in ssRNA infections of Escherichia coli, OCb5 can be released from infected cells of C. crescentus without cell lysis, and infected cells can generate macroscopic colonies. To determine the mechanism by which infected cells release naked, spherical virions without lysis, ultrathin sections of infected cells will be examined with particular attention to changes in the membranes, sacculus and periplasm of the cell surface. ssRNA phages have been isolated for two other genera, Pseudomonas and Escherichia, and infected cultures are known to become "temporarily immune" to infection. To determine whether the observations of OCb5 infection of C. crescentus explain the mildness of infection in other ssRNA phage-host systems, infected cultures of these other organisms, as well as of other species of Caulobacter, will be tested for secreted ribonuclease and for bIFN. The overall goal of the research is to determine whether prokaryotes, like animal cells, can employ a (polypeptide?) signal that confers an anti-viral state. If so, this capability could account for the persistent coexistence of bacteria and their viruses, as well as provide a prokaryotic model for a biologic relationship whose understanding could assist in the control of viral infections of humans.